Variable Inclination Centrifugation Assembly for Rapid Separation of Blood

This invention relates to instruments and methods for separating whole blood into its components.

Most clinical chemistry analysis requires a separated blood sample. Separation of a usable sample from whole blood is traditionally achieved by centrifuging blood collection tubes for several minutes. The current practice is for tubes be spun for approximately 10 to 15 minutes at 1200 g.

The current method has several drawbacks, however. First, the blood separation time delays delivery of analytical results. Second, the long separation time requires large centrifuge batch sizes in order to maintain high sample throughput during blood processing. This necessitates the use of large, expensive centrifuges.

Rapid blood separation systems have been developed. For example, see U.S. Pat. Nos. 5,030,341 and 5,308,506 to McEwen et al. These patents describe specialized blood separation tubes and a specialized instrument for rotating the tubes about a longitudinal axis. U.S. Pat. Nos. 4,846,974 and 4,981,585 to Kelley et al. disclose disk-shaped cassettes for centrifugal separation of blood with a high speed rotary drive. All of the above systems require specialized blood collection or separation containers in addition to dedicated instruments, which add a continuing burden to the overall cost of the system.

What is needed is a blood separation system and method which is easy to use, contributes to the low cost operations of the laboratory, and rapidly provides a separated blood sample.

The above objects have been achieved with a variable inclination centrifugation assembly and method for rapid separation of samples, such as blood, which have components of differing densities. In a first phase, the samples, which are contained within tubes and tube holders, are rotated about a central axis of a yoke which supports the tube holders, while the tube holders are in a first position in which the longitudinal axes of the tube holders are substantially parallel to the axis of rotation. In a second phase, the tube holders are carefully and gradually inclined outwardly or radially, while rotation continues, toward a second position in which the longitudinal axes of the tube holders are substantially perpendicular to the rotation axis.

The method of the present invention allows for rapid separation because spinning of the sample initially, i.e. in the first phase, allows the sample to travel a distance equal to less than the diameter of the tube. This causes preliminary separation of the sample components. As the tube holders are inclined, the separation of sample components is refined and stabilized as the sedimented sample components travel toward the bottom of the tube. The decrease in separation time may be twelve-fold over conventional methods.

The first embodiment of the variable inclination centrifugation assembly includes a rotation subassembly and an inclination subassembly preferably utilizing a worm gear. A separate rotation motor and inclination motor are utilized, and the rotation and inclination aspects of the separation may be independently controlled. Because the worm gear is engaged, the tube holders do not swing freely, but rather become inclined upon operation of the inclination subassembly.

The second embodiment of the centrifugation assembly utilizes a spring-loaded linkage system in which the spring-loaded linkages are activated by the rotational speed of the samples and cause the inclination. Each tube holder is coupled via one spring-loaded linkage to an anchor ring that is positioned about the central hub of the centrifugation assembly. Housed within each linkage are a compression spring and an extension rod. At rest and during low speed rotation, the tube holders are each held in the first position by the associated spring-loaded linkage, whose overall length is dependent upon the force exerted by its internal compression spring. As the speed of rotation increases, the forces arising from centrifugation overcome the spring force within the linkage and allow the extension rod to compress the spring and thus extend the length of the spring-loaded linkage. This action of the spring-loaded linkages causes the tube holders, which are connected to the yoke at pivot points, to become inclined toward the second position. Through the use of appropriately-sized compression springs, the inclination occurs gradually, so as not to disrupt the sample.

The third embodiment includes a governor-based linkage system. An adjustable governor mass in combination with a set of three arms extending from the mass is associated with each of the tube holders in this embodiment. One of the three arms of each governor-based linkage is connected to one of the tube holders, another arm is connected to the yoke or hub of the centrifugation assembly, and a third arm is connected to a universal collar that slides about the hub of the centrifugation assembly. A resistance spring is also positioned about the hub and presses down upon the sliding collar, thereby retaining the tube holders in the first position at rest and at low speeds. As rotational velocity increases, however, each governor mass is driven outward from the hub. This outward drive compresses the resistance spring to establish an equilibrium position for each speed. Each governor mass with its associated three arms moves in an outward direction from the hub, thereby causing pivoting of the tube holders and inclination of the samples within the tube holders. As with the second embodiment, an appropriately-sized spring may be used to ensure gradual and synchronized inclination of the samples and to maintain balance within the assembly.

An advantage of the present invention is that separation of the samples occurs rapidly and, in the case of blood samples, conventional blood draw tubes may be used. Additionally, the assemblies need not be fixed parts of the centrifuge. Therefore, they may be utilized in conjunction with standard centrifuges and may be removed from the centrifuge for sterilization temperature control, or remote sample preparation.

FIG. 1 illustrates the effects of the motions and relative positions of the tubes upon the samples.

FIG. 2 is a side view of the centrifugation assembly according to the first embodiment of the present invention, with a sectional view of the hub.

FIG. 3 is a detailed view of a section of FIG. 2, showing the worm gear of the first embodiment.

FIG. 4 is a top view of the centrifugation assembly of FIG. 2.

FIG. 5 is a side view of a portion of the centrifugation assembly according to the second embodiment of the present invention, showing a spring-loaded linkage and a tube holder in a first position.

FIG. 6 shows a second position of the tube holder of FIG. 5 and the spring-loaded linkage in an extended position.

FIG. 7 is a sectional view of the spring-loaded linkage of FIGS. 5 and 6.

FIG. 8 is a side view of a portion of the centrifugation assembly according to the third embodiment of the present invention, showing a governor-based linkage and a tube holder in a first position.

FIG. 9 shows a second position of the tube holder of FIG. 8 and the governor-based linkage in a radially extended position.

FIG. 10 is a perspective view with a partial cutaway of the governor mass of FIGS. 8 and 9.

With reference to FIG. 1, the relative positions of a sample within a tube, according to the method of the present invention, is shown. Balanced tubes are initially held in a first position in which the longitudinal axes of the tubes are substantially parallel to the rotation axis 61. The tubes are rotated about axis 61, as shown in FIG. 1 by arrow "A." Rotation while the tubes are in this first position causes the denser elements of the sample, such as red blood cells in whole blood, to settle along the outer walls of the tubes. The time required for the settling is short compared to conventional centrifugation, as the cells need to travel only about half the tube diameter. Cells in tubes spun conventionally, by contrast, must settle about half the length of the tube.

Preferably after the majority of cells have settled, the tubes are inclined with respect to the rotation axis 61. The result is that the angle between the longitudinal axis of each tube and the rotation axis 61 increases from near zero to near ninety degrees, as shown by arrows "B" in FIG. 1. Thus, the longitudinal axes of the tubes and the rotation axis 61 begin in a substantially parallel position and are shifted to a substantially perpendicular position with respect to each other. Rotation continues during this inclination change. The purpose of the inclination change is to shift the separated layers of blood or other sample from an arrangement which is stable only under rotation to one which is stable without rotation. To accomplish this, the inclination change should be gradual. An abrupt tilt may remix the pellet with the supernatant.

After the inclination change is complete, the tube rotation may stop. For a blood sample, this leaves the cells at the bottom of the tube and supernatant, e.g. serum or plasma, above. It is desirable to restore the tube to the first position for ease of handling. This can be done after the rotation has stopped or as the rotation is stopping. If the latter method is chosen, then it is important that the rotation be sufficiently slow during this restoration so that the sample components do not reorient to the arrangement achieved during the earlier phase. An appropriate rotation speed during this restoration is one where the resultant centrifugal force is less than the gravitational force. The nature of the sample dictates the degree of inclination and the top speed of rotation necessary for the separation process.

In FIG. 1, tubes 11 containing samples of whole blood 30 are shown. During the first phase, the samples generally travel toward the outer walls of the tubes. The second phase allows for further separation into a serum or plasma supernatant 32 and a pellet of blood cells 31.

For the initial phase, the tubes need not be oriented exactly parallel to the rotation axis. A slight positive inclination, with the bottoms of the tubes further from the rotation axis than the tops of the tubes, reduces or eliminates hydrodynamic forces on the tube stoppers 64. This may be desirable to prevent ejection of the stopper and loss of the sample. Also, the assembly need not be oriented so that the rotation axis is vertical.

FIG. 2 shows a centrifugation assembly of the present invention, according to a first embodiment. This first embodiment allows for independent control of the rotation and inclination in the centrifugation process, and is the preferred embodiment. In FIG. 2, tube holders 12, which hold tubes 11, are shown suspended from a radially-extending rotor yoke 15. The tube holders 12 are spaced apart from and balanced about the central axis of yoke 15. Balancing of the tube holders is generally accomplished by positioning tube holders with contents of equal mass symmetrically about the central axis. A tube holder may be balanced with a weight, however. For clarity, FIG. 2 shows only two tube holders and associated supports. The tube holders 12 are connected to the yoke 15 by pivots 13 and bearings 63. Thus, tube holder 12 may move about pivot 13.

The rotor yoke 15 may be connected to the rotor 26 via rotor extensions 60. Rotor 26, according to the first embodiment, is a hollow cylinder with projecting extensions 60 for support of yoke 15. Rotor 26 is driven by rotation motor 23 which connects to pulley 25 via armature shaft 57. Belt 21 surrounds rotor 26 and connects pulley 25 to rotor 26. The rotation motor 23 therefore drives the pulley 25 and belt 21 assembly and thereby drives the rotor 26 to cause rotation about the rotation axis. The rotation motor 23 thus controls the first phase of centrifugation according to the present invention.

Also within the centrifugation assembly of the first embodiment is a shaft 20, which is driven to rotate about its central axis by inclination motor 24. About one end of the shaft 20 is a worm portion 16 of a worm gear. The worm 16 is in engagement with the teeth of various worm wheel segments 14. Each worm wheel segment 14 is operably connected to one of the tube holders 12, preferably through ring piece 17 and its extension 19, as seen in FIGS. 2 and 4. Rotation of shaft 20 by inclination motor 24 causes rotation of worm 16. In turn, rotation of worm 16 causes worm wheel segments 14 to travel relative to worm 16. Since the worm wheel segments 14 are fixedly attached to tube holders 12, movement of the worm wheel segments 14 causes movement of the tube holders 12 by pivoting of tube holders 12 about pivots 13. A cone drive gear may be an appropriate alternative to the worm gear.

FIG. 3 presents an enlarged view of the worm gear operation, according to the first embodiment. In FIG. 3, teeth 22 of worm wheel segment 14 are clearly visible. By rotation of shaft 20 in a given direction and corresponding rotation of the worm in that direction, movement of the worm wheel segments 14 relative to the worm 16 occurs along the top halves of the worm wheel segments. Thus, tube holders 12 move about pivots 13, and are caused to incline with their top ends closer to the central or rotation axis of the centrifugation assembly than their bottom ends. Shaft 20 is physically located within rotor 26, as seen in FIG. 2, but the movement of shaft 20 is independent of the movement of rotor 26. Inclination motor 24 thus controls the second phase of centrifugation according to the present invention.

Rotor 26, shaft 20, and spindle 27 constitute the hub of the centrifugation assembly of the first embodiment, and may be supported on base 58 of FIG. 2. Spindle 27 is a fixed portion of the hub and serves to separate shaft 20 from rotor 26. Base 58 may serve to anchor spindle 27. Spindle 27 has an annular cross section, and in its simplest embodiment is a hollow cylinder. Shaft 20 is housed within spindle 27. Shaft 20 and spindle 27 are separated by bearings 28, which are preferably ball bearings. These bearings 28 allow shaft 20 to rotate freely within fixed spindle 27. Spindle 27 is itself positioned within rotor 26. Shaft 20, spindle 27, and rotor 26 have a common longitudinal axis. This is also coaxial with the central axis of yoke 15. As shown in FIG. 2, bearings 29 are positioned between spindle 27 and rotor 26. As with the bearings 28 between spindle 27 and shaft 20, the bearings 29 between spindle 27 and rotor 26 are preferably ball bearings. Rotor 26 is thus permitted to rotate freely about spindle 27. Because spindle 27 is fixed and separates the motion of shaft 20 from the motion of rotor 26, and because shaft 20 and rotor 26 are driven by independent motors, the centrifugation assembly of the first embodiment allows for independent control of rotation and inclination.

In operation, shaft 20 and rotor 26 should rotate synchronously to maintain a fixed inclination. If shaft 20 turns faster than rotor 26, the angle of inclination increases. If shaft 20 turns slower than rotor 26, the angle of inclination decreases. The high gear ratio available from the worm gear permits the limited torque of the inclination motor to maintain a fixed inclination at the high loads produced by rapid rotor rotation.

Unlike centrifuges of the prior art, for example, see U.S. Pat. No. 4,221,324 to Frey, the centrifugation assembly of the present invention does not include freely-swinging tubes or tube holders. Instead, inclination may be carefully controlled, both during rotation and, in the first embodiment, independent of rotation. Because the worm 16 is engaged with worm wheel segment 14 in the first embodiment, tube holders 12 may not freely swing about pivot 13. In the second and third embodiments of the centrifugation assembly of the present invention, strong springs may be utilized to control the rate and degree of inclination of the samples and prevent the tube holders from swinging freely about their pivots.

With reference to FIG. 4, rotor yoke 15 is shown as being capable of supporting multiple tube holders 12. FIG. 4 shows a cross-shaped yoke 15 capable of supporting four samples. In FIG. 2, two tube holders were shown for the sake of clarity. It is understood that rotor yoke 15 may be adapted for supporting two, four, or any other convenient multiple of tube holders. Additionally, yoke 15 may be adapted for supporting easily replaceable tube holders of varying shapes and sizes, including bucket-type holders for large sample containers. The tube holders may each hold more than one tube or sample.

FIG. 4 shows the top of shaft 20 having worm 16, and engagement of worm 16 with the worm wheel segments 14, one worm wheel segment 14 for each of the sample positions of the assembly. Worm wheel segments 14 are preferably semicircular plates having teeth along the outer periphery of the arced portion. The straight, non-toothed, portions of the worm wheel segments 14 are fixedly attached to tube holders 12. In FIG. 4, it is shown that this is advantageously accomplished through the use of ring pieces 17, wherein each ring piece 17 has an extension 19 for fixed attachment of a worm wheel segment 14. Each ring piece 17 may also serve as the support point for a pivot 13. For example, in FIG. 4, each pivot 13 is shown as two pieces 13a and 13b connecting ring piece 17 to yoke 15, comprising respective end pieces of a pivot shaft, and allowing movement of tube holder 12 about pivot axis 13c. Bearings 63, preferably ball bearings, may be used to support pivot 13 within yoke 15. Ring piece 17 may have tube holder 12 placed or attached within it. Alternatively, ring piece 17 may be an integral part of tube holder 12. A rim 18 is shown in the embodiment of FIG. 4 as connecting the various extensions of rotor yoke 15. This rim is not necessary, but may be desired for increased stability.

Referring to FIGS. 5 and 6, the centrifugation assembly according to the second embodiment of the present invention is shown. This second embodiment utilizes a spring-loaded linkage 33 to effect inclination, instead of a worm gear. In FIG. 5, the hub 35 of the centrifugation assembly is attached to rotor yoke 15. Hub 35 and yoke 15 may be of simpler construction in the second and third embodiments than in the first embodiment. For example, the hub of FIG. 5 may be a solid piece. As in the first embodiment, hub 35 is coaxial with the central axis of yoke 15. Tube holder 12 is shown supported from yoke 15 at pivot 13, as before. In the spring-loaded linkage system, an anchor ring 37 is shown disposed about hub 35. Spring-loaded linkage 33 is attached at one end to anchor ring 37 and an opposite end to tube holder 12, as by hinge piece 34 which is attached to tube holder 12. Alternatively, anchor ring 37 may be eliminated and the first end of each spring-loaded linkage 33 may be attached directly to hub 35. However, the anchor ring provides an advantage in that it may be fixed about hub 35 during processing of the samples, but may be moved and fixed at a higher or lower position along hub 35 for different sample runs, thereby affecting the inclination rate. Within a cylindrical housing 41 of spring-loaded linkage 33, as seen in FIG. 7, there are a compression spring 40 and an extension rod 36. The extension rod 36 is attached to a compression disk 39 within the housing 41 at one end. The other end of extension rod 36 extends from the housing 41 and is the point at which the spring-loaded linkage 33 is attached to anchor ring 37. Compression spring 40, compression disk 39, and a substantial portion of extension rod 36 are fitted within the cylindrical housing 41 of the spring-loaded linkage 33 by threaded adjustable bearing 38. Threaded adjustable bearing 38 centers extension rod 36 and adjusts the tension within housing 41 and thereby adjusts the rate of inclination of the tube holder 12.

At rest and during low speed rotation, as by rotation means 62, in the second embodiment of the centrifugation assembly, the tube holder 12 is held in the first position by the spring force of the spring-loaded linkage. The length of the spring-loaded linkage 33 is controlled by the force exerted by compression spring 40. As the speed of rotation of the samples increases, the forces arising from centrifugation overcome the spring force and allow extension rod 36 in combination with compression disk 39 to compress compression spring 40. This compression results in an extension of the length of spring-loaded linkage 33.

With reference to FIG. 7, compression of spring 40 and corresponding extension of extension rod 36 out of the housing 41 occurs in direction "C." Relative motion of the cylindrical housing 41 of spring-loaded linkage 33 occurs in direction "D."

FIG. 5 therefore represents centrifugation during the first phase of operation wherein the tube holders are held in the first position. FIG. 6 represents the second phase, wherein the tubes are inclined toward the second position, because centrifugal forces have overcome the spring force of compression spring 40 and allowed extension of spring-loaded linkage 33. Through the use of a spring having a high spring force, and through bearing 38, inclination can be controlled so that it occurs gradually. The length of the housing 41 and extension rod 36 should also be sized to allow smooth, kinematic movement of tube holder 12.

FIGS. 8 and 9, present a portion of the centrifugation assembly according to a third embodiment of the present invention. Other than the linkage system, the third embodiment is similar to the second embodiment. This third embodiment utilizes a governor-based linkage system to effect inclination. In particular, a governor-based linkage 42 includes a governor mass 44 and a set of three linking arms. The first arm 46a connects governor mass 44 to the tube holder 12, as by hinge piece 34. The second arm 46b is attached to the hub 35 or yoke 15 of the centrifugation assembly. The third arm 46c is attached to a sliding collar 47. Sliding collar 47 is disposed about hub 35. Also disposed about hub 35 is resistance spring 43.

In operation, resistance spring 43 pushes sliding collar 47 downward on hub 35 through its spring force, thereby retaining the tube holder 12 in the first position. As rotational velocity increases, however, the governor mass 44 is driven outward or radially from hub 35. This outward drive compresses resistance spring 43 in order to establish an equilibrium position for each speed. The force compressing the spring depends upon the mass and upon the radial position of the mass. The result is a first phase of centrifugation with tube holder 12 in the first position, as shown in FIG. 8. Tube holder 12 then moves toward the second position, as shown in FIG. 9, as the rotational velocity of the samples increases. Once again, through the use of a spring having a high spring force, inclination can be caused to occur gradually.

The sliding collar 47 also serves to synchronize the inclination rate of the various pivoting tube holders. This is important for dynamic balance of the assembly and smooth movement of the tube holders. Dynamic imbalance may result in ruined samples and a damaged assembly or centrifuge.

FIG. 10 is a detailed partial cutaway view of the preferred governor mass 44 of the third embodiment of the present invention. Particularly, governor mass 44 includes a slotted cylindrical housing 52 which houses an adjustment mass 48 and a restoring spring 49. The radial position of the adjustment mass 48 is adjustable by turning adjusting screw 50 which moves adjustment mass 48 against restoring spring 49 and thereby adjusts the rate of inclination of the connected tube holder. At least one pin 54 mates adjustment mass 48 with a slot 53 in the cylindrical housing 52 to prevent rotation of adjustment mass 48 about the screw axis. The housing is closed by a threaded cap 59. The outside of cylindrical housing 52 is provided with arm link extensions 51a-c for connecting governor mass 44 with each of the three linking arms 46a-c.

Although only one tube holder 12 and one spring-loaded linkage 33 or one governor-based linkage 42 are depicted and described in the second and third embodiments respectively, each tube holder 12 supported on yoke 15 of the centrifugation assembly has an associated linkage. These linkages serve as the spring means for controlling the inclination of the samples during centrifugation. Rotation in these embodiments is via a simple rotation motor 62, shown in FIGS. 5 and 8, or by the rotation drive of the centrifuge into which the assemblies are installed.

Although an even number of tube holders positioned symmetrically about a yoke represents the preferred configuration for balance purposes in the three embodiments, a singular tube holder, or other odd number of tube holders, may be supported from the yoke of the centrifugation assembly. This odd-numbered configuration necessitates a properly-positioned counterbalance, however.

As stated above, the centrifugation assembly of the present invention need not be a fixed part of a centrifuge. This allows the assembly to be used in various standard centrifuges. Since standard blood draw tubes of glass or plastic may be utilized with the assembly, the result is a one-time cost of the assembly to laboratories and hospitals. The assembly may also be removed from the centrifuge for cleaning, sterilization, temperature control, or sample preparation in a protected environment.

Depending upon the details of the centrifuge that will incorporate the apparatus of the present invention, the three embodiments of the assembly may include a rotation motor or other means for rotating the samples about the central axis, as previously shown and described, or the assemblies may be rotor assemblies attachable to an existing rotation motor of the centrifuge. The unusual position of the rotation motor in the first embodiment, as compared to most standard centrifuges, generally indicates that an assembly with a rotation motor included and positioned as shown in FIG. 2 is likely to be the more useful variation of the first embodiment, however.

The three embodiments of the centrifugation or rotor assembly and the method for rapid separation of blood described herein represent a simple, yet significant, solution to the problems of high throughput blood processing.